Transcriptional and Posttranscriptional Regulation of Cyclooxygenase-2 Expression by Fluid Shear Stress in Vascular Endothelial Cells

Abstract

Objective-Fluid shear stress induces cyclooxygenase (COX)-2 gene expression in vascular endothelial cells. We investigated the underlying mechanism of this induction. Methods and Results-Exposure of human umbilical vein endothelial cells to laminar shear stress in the physiological range (1 to 30 dyne/cm 2 ) upregulated the expression of COX-2 but not COX-1, a constitutive isozyme of COX. The expression of COX-2 mRNA began to increase within 0.5 hour after the loading of shear stress and reached a maximal level at 4 hours. Roles of the promoter region and the 3Ј-untranslated region in the human COX-2 gene were evaluated by the transient transfection of luciferase reporter vectors into bovine arterial endothelial cells. Shear stress elevated luciferase activity via the region between Ϫ327 and 59 bp. Mutation analysis indicated that cAMP-responsive element (Ϫ59/Ϫ53 bp) was mainly involved in this response. On the other hand, shear stress selectively stabilized COX-2 mRNA. Moreover, shear stress elevated luciferase activity when a 3Ј-untranslated region of COX-2 gene containing 17 copies of the AUUUA mRNA instability motif was inserted into the vector. Key Words: shear stress Ⅲ vascular endothelial cells Ⅲ cyclooxygenase-2 Ⅲ posttranscriptional regulation V ascular endothelial cells are always exposed to a wide variety of biochemical and biomechanical stimuli, including fluid shear stress caused by blood flow. Shear stress modulates several endothelial functions, such as control of vascular tone, maintenance of antithrombotic surfaces, regulation of inflammation, protection against oxidative stresses, and regulation of endothelial cell proliferation and apoptosis. 1 Cyclooxygenase (COX), a rate-limiting enzyme for prostaglandin (PG) biosynthesis, comprises 2 isozymes, COX-1 and COX-2. 2,3 COX-1 is constitutively expressed in most cell species, whereas COX-2 is an inducible enzyme whose expression is regulated differently among cell types. Growing evidence indicates that COX-2 plays a key role in several biological processes, such as inflammation, tumorigenesis, development, and atherogenesis. 4 -9 Laminar shear stress upregulates COX-2 gene expression. 10 COX-2 is involved in lipopolysaccharide-stimulated production of prostacyclin (PGI 2 ) in endothelial cells 11 and is also involved in PGI 2 biosynthesis in healthy humans. 12 Previously, we have reported that shear stress promotes the production of PGD 2 in endothelial cells by stimulating the expression of lipocalintype PGD 2 synthase (L-PGDS), whereas PGI 2 synthase was constitutively expressed but did not respond to shear stress. 13 Therefore, the induction of COX-2 expression by shear stress may be involved in the production of PGI 2 and PGD 2 in endothelial cells. Conclusions-TranscriptionalThree cis-acting elements, namely, the nuclear factor (NF)-B binding site, the NF-interleukin-6 (NF-IL6) binding site, and the cAMP-responsive element (CRE), reside in the region between base pairs Ϫ327 and ϩ59 (Ϫ327/ϩ59) in the human COX-2 gene promoter. Their involveme...

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